Borrelia burgdorferi, the agent of Lyme disease, is an obligate parasite of its mammalian reservoirs and of the tick vectors that transmit the bacteria. B. burgdorferi infection can affect the skin, joints, heart, and nervous system, and can withstand the onslaught of the innate and acquired immune systems. Interactions with mammalian cells are likely to be important in this complex infection process, and B. burgdorferi binds to three members of the integrin family of mammalian cell-surface receptors. The B. burgdorferi protein p66 is a ligand for integrins alpha IIb beta3 and alpha/v beta3, but does not bind integrin alpha5beta1, suggesting that other ligands are expressed by the spirochete. P66 is expressed by B. burgdorferi in the mammal, but not in the tick, supporting its potential importance in mammalian infection. To further study the expression of p66, the importance of p66 in the biology of mammalian infection by Borrelia, and to identify additional B. burgdorferi proteins that bind to integrins, three Specific Aims will be pursued: Aim 1. In vitro and in vivo analysis of B. burgdorferi mutants that do not express p66. P66-deficient B. burgdorferi will be compared to wild type with regard to growth and ability to establish infection. Aim 2. Analysis of the regulation of B. burgdorferi p66 expression. The pattern of p66 expression suggests that it is a paradigm for B. burgdorferi genes that are important in the mammalian host, so genetic and biochemical approaches will be taken to identify B. burgdorferi modulators of p66 expression. Aim 3. Characterization of potential integrin ligands identified by analysis of the B. burgdorferi genome sequence. Five B. burgdorferi proteins that contain integrin-recognition motifs and secretion signals will be tested for integrin-binding, localization on the bacterial surface, and for roles in infection. These experiments will shed light on the importance of p66 and integrin attachment in the ability of B. burgdorferi to cause infection, and on regulation of gene expression in this important pathogen. The long-term goal of this work is to understand the pathogenesis of B. burgdorferi infection, which is an important model for how certain bacteria co-opt the biology of their immunocompetent hosts to cause disease.